Multiple ephrins regulate hippocampal neurite outgrowth.

Increasing evidence indicates that the eph family of ligands and receptors guides the formation of topographic maps in the brain through repulsive interactions. For example, we have recently found that in the hippocamposeptal system, the ligand ephrin-A2, which is expressed in an increasing gradient from dorsal to ventral septum, selectively induces pruning of topographically inappropriate medial hippocampal axons. The recent detection of ephrins A3 and A5, as well as A2, in the septum raised critical functional questions. Do the ligands act combinatorially, ensuring appropriate three-dimensional spatiotemporal projection, or do they exert entirely distinct actions in addition to guidance mechanisms? To approach these alternatives, we cloned mouse ephrin-A2 and compared the activities of the three ligands. Here, we show that these ligands reduce the number of hippocampal neurites in a similar fashion. The effect was regionally specific; medial hippocampal neurites were reduced 1.5- to 1.8-fold, whereas lateral hippocampal neurites were not significantly affected, conforming to topographic projection in vivo. Furthermore, we found that ephrins regulated neurite number in a stage-specific fashion, affecting E19 hippocampal neurites more than E16 neurites. Our observations suggest that all three septal ephrins, A2, A3, and A5, play spatiotemporally specific roles in guiding topographic projections from the hippocampus.